The giant synapse of the hatchetfish presents unique possibilities for investigation of synaptic transmission. Both pre- and postsynaptic elements can be penetrated by independent microelectrodes and postsynaptic resistance is large enough that effects of single quanta can be recorded. Transmission is cholinergic. Tetanic stimulation rapidly depletes the synapse of vesicles PSP amplitude to a point where release of incompletely filled vesicles is indicated. Filling of vesicles after depleting tetanus is a rapid process and we propose to follow the time course using ramp stimuli that cause asynchronous release. Treatment with hemicholinium is expected to increase filling time. Tetanic stimulation causes increase in the amount of presynatptic membrane in cysternae. We plan to use extracellular tracers such as horseradish peroxidase to follow the presumed recycling of membrane from vesicles to surface to coated vesicles to cysternae to vesicles again. We will make inventories of total membranes in the several stages under conditions of stimulation and recovery at room and low temperature. The number of vesicles will be correlated with the mobilizable store of quanta. The effects of divalent ions will be studied with intracellular as well as extracellular application. We should be able to separate relative activity of divalent ions at intracellular release sites from relative permeabilities through the surface membranes. Intracellular injection of enzymes, metabolites or toxins should allow further analysis of processes of transmitter synthesis packaging and release.